The present invention relates to embedding media for microscopy specimens and more particularly to a low viscosity embedding medium for numerous types of electron microscopy specimens.
In sectioning specimens for subsequent microscopic examination it is desirable to secure sections with the least amount of distortion or disruption of cell or tissue structures as they existed in the viable or original state. In order to preserve such structures it is the general practice to infuse the specimen before sectioning with a medium designed to firm up or reinforce such microstructures so that they will remain intact during the sectioning process and during subsequent handling while under examination.
Such embedding media range from the old paraffinic materials to more recently devised organic polymeric materials such as acrylics and epoxy compositions. Despite the improvement in composition of embedding media a continuing problem remains, i.e., complete infusion of the microstructures. For unless the embedding medium thoroughly penetrates into the minutest structure, distortion or disruption thereof will take place upon sectioning and subsequent handling.
Thorough infusion of the specimens is also complicated by the fact that while the medium must completely penetrate the specimen microstructures, it must also be resistant to fracturing upon sectioning, it must remain transparent to light or the electron beam, it must show no microstructure itself at the microscopy magnifications to be utilized, it must solidify or polymerize at temperatures low enough to prevent damage to the specimen structure or tissues, it must not damage the specimen microstructures upon polymerization, but it must solidify or polymerize to sufficient hardness to rigidly hold the specimen microstructures in position during sectioning and subsequent handling, and finally it must possess a reasonable "pot" life to permit infusion of a reasonable number of specimens or large specimen samples before solidification or polymerization takes place.
A new embedding medium has now been devised which fulfills all of the aforestated requirements as well as being of sufficiently low viscosity to thoroughly penetrate and infuse the minutest structures of a wide range of specimen tissues including problem materials such as the lipid storage endosperm of castor bean seeds and tomatoes, hard, lignified elements in vascular bundles of tomato leaves, as well as the soft, highly vacuolated, parenchymatous tissues of ripe tomato fruits.
Briefly the medium of the invention comprises low viscosity components comprising the following: a cycloaliphatic diepoxide, preferably vinyl cyclohexene dioxide; an epoxy resin flexibilizer, preferably diglycidyl ether of polypropylene glycol; an anhydride hardener, preferably nonenyl succinic anhydride; and an accelerator, preferably an alkyl alkanol amine such as dimethylamino-ethanol.